Effect Of Astragalus Polysaccharide (APS) On The Activity Of AMPK Involved In Improvement Of Glucose Metabolism In Type 2 Diabetes Mellitus

Type 2 diabetes mellitus (T2DM) is the major form of diabetes mellitus and is associated with pancreaticβcells function disorder, insulin resistance, obesity, hyperglycemia, hyperlipidemia, and chronic inflammation, among other symptoms. Hyperglycemia is the basic pathophysiological characteristic of T2DM,which is the main root of the malfunction of organ and the various subsequent complications. In the case of T2DM, protection of pancreatic P cells function, augmentation of glucose utilization have been strategies taken to improve the glucose metabolism associated with diabetes, In addition to the classical insulin signal pathway, the glucose uptake is also regulated by AMPK signal pathway which acts essentially like a "fuel gauge". AMPK activity can also regulate lipid oxidation and synthesis and now become a potential target for therapeutic treatment of T2DM.Astragalus polysaccharide (APS) is the polysaccharide component of the ethanol extract of Astragalus roots and is the active component of Astragalus. In our previous study, we found that treatment with APS lowers blood sugar by decreasing the expression and activity of PTP1B in the skeletal muscles of T2DM animals. We also found that APS can improve insulin sensitivity, which is coupled with an enhanced adaptive capacity of the endoplasmic reticulum.The hypoglycemic effects and improvement of glucose metabolism of APS on T2DM is more likely the effects of AMPK. What we concerned is whether APS can develop the hypoglycemic effects and improvement of glucose metabolism via AMPK pathway.T2DM model has been established to investigate whether APS plays a direct role in improving glucose metabolism in vivo. We use pancreas、liver and skeletal muscle to detect the function of pancreaticβcells、glycogen synthesis and skeletal muscle glucose translocation. In addition, we also investigated whether the hypoglycemic effects of APS involve the AMPK signal pathway in vivo. We sought to explore the possible mechanism of APS involved in the regulation of glucose metabolism. Our experiments are composed of two main parts, in vivo and in vitro.Part I Effect of APS on the activity of AMPK involved in improvement of glucose metabolism in type 2 diabetes mellitus In this study, T2DM rat model was duplicated by a high-fat diet (58%fat,25.6% carbohydrate, and 16.4%protein) and a small dose of streptozotocin (STZ,25mg/kg, ip). After APS therapy (700mg/kg/d, ig) for 8 weeks, blood glucose, glycosylated hemoglobin, and serum insulin were measured. Insulin sensitivity was evaluated by the comprehensive analysis of OGTT and HOMA IR index. And more importantly we want to observe the hypoglycemic effect of APS and its potential regulation of pancreatic (3 cells function in pancreas、glycogen synthesis in liver and glucose translocation in skeletal muscle. Great attentions are put on the expression and activity of AMPKa in skeletal muscle, a novel activators of new strategy for the T2DM treatment. This study is designed to investigate underlying the regulation of glucose metabolism by APS.The results showed that we have successfully established the T2DM model which has some typical characteristics as follows:hyperglycemia, high postprandial blood glucose, and high glycosylated hemoglobin, but had normal levels of insulin compared with normal rats; a islet amyloidosis withβcells apoptosis in pancreas; a significant reduction in hepatic glycogen content, AMPK activity of skeletal muscle and the expression of GLUT4 also in the plasma membrane extract and the cytoplasm extract.The hyperglycemia status, insulin sensitivity, function of pancreatic (3 cells, glucose uptake, and activation level of AMPK in T2DM rats were improved in response to APS administration. We could speculate that APS can improve glucose metabolism by alleviating pancreatic islet amyloidosis with protectingβcells from being destroyed and apoptosis, increasing liver glycogen synthesis and skeletal muscle glucose translocation in the T2DM rat model, via activation of AMPK.Part II Mechanism of APS on the activity of AMPK involved in improvement of glucose metabolism in type 2 diabetes mellitusThere are two major mechanisms for increasing glucose transport in skeletal muscle. One is the insulin signaling pathway, by which activation of PI3K/Akt phosphorylation leads to GLUT4 translocation to the plasma membrane. The other is the AMPK signaling pathway. Increasing evidence suggests that the activation of AMPK promotes glucose uptake via a PI3K-independent pathway. On account of the study of PartⅠ, we want to find the "bridge" which linked APS and glucose uptake, to establish whether APS can active AMPK and increase glucose uptake in vitro, and more important is, whether AMPK signaling pathway or insulin signaling pathway. So we add Wortmannin which is the inhibitor of PI3K-insulin pathway and Compound.C which is the inhibitor of AMPK pathway in our experiments.The results showed that the activity of AMPK was increased by an increased concentration of APS (50-200μg/ml) in C2C12 mouse skeletal muscle cells. APS treatment can also increased glucose uptake significantly in C2C12 cells. Compound C, which is a highly-selective AMPK inhibitor, could effectively block APS-induced glucose uptake. However, wortmannin, the PI3K signaling pathway inhibitor, had less effect on APS-induced glucose uptake, indicating involvement of the AMPK pathway in these APS effects.The experiments in vitro also showed that high glucose treatment of C2C12 cells induced a reduction in AMPK activity. Treatment with APS increased AMPK activity and glucose uptake. However, this effect could also be suppressed by Compound C. APS could improve glucose metabolism through increases in the AMPK pathway activity in C2C12 cell model.Conclusion:1. Rat model of T2DM was successfully established. The hyperglycemia status, insulin sensitivity in T2DM rats were improved in response to APS administration.2. APS can regulate glucose metabolism at least in part by alleviating islet amyloidosis with protectingβcells from being destroyed and apoptosis, increasing hepatic glycogen content, skeletal muscle AMPK activity and GLUT4 expression also in plasma membrane extract and cytoplasm extract.3. APS can increase glucose uptake in C2C12 cells via AMPK-dependent pathway. APS can also increase AMPK activity and glucose uptake in high glucose treatment of C2C12 cells. APS can regulate glucose metabolism in vitro.